The lipin proteins (lipin-1, lipin-2, and lipin-3) are critical for triacylglycerol (TAG) metabolism, a key theme of our Program Project. Lipin proteins have phosphatidate phosphatase (PAP) enzyme activity and catalyze the penultimate step in TAG synthesis, converting phosphatidic acid to diacylglycerol. Lipins also interact with transcriptional regulators to modulate gene expression. Lipin-1 expression levels and activity influence adiposity, insulin resistance, liver and muscle lipid homeostasis, energy expenditure, and peripheral nerve myelination. The physiological roles of lipin-2 and lipin-3 are much less well characterized. Human lipin-2 deficiency causes Majeed Syndrome, which is distinguished by bone inflammation and anemia. The mechanism for this disease is unknown and highlights the need for a better understanding ofthe physiology and pathophysiology of lipin proteins. Virtually nothing is known about the physiological role of lipin-3. Our proposal is based on three discoveries that we have made regarding the biological roles of lipin-2 and iipin-3 using our lipin-deficient mouse models. The first is that lipin-2 knockout mice have abnormalities in the bone growth plate, and lipin-2 is localized to cells that populate the growth plate?the chondrocytes. In Aim 1 we will define the role of lipin-2 in chondrocyte differentiation and determine the mechanisms underlying bone abnormalities associated with lipin-2 deficiency. The findings may provide fresh insights into the pathogenesis of bone abnormalities in Majeed syndrome. A second discovery is that adipose tissue PAP activity and TAG accumulation are determined not only by lipin-1, but also by lipin-3. We have mapped a physical interaction between lipin-1 and lipin-3, suggesting a mechanistic basis forthe requirement of both lipins in adipose tissue. Our findings regarding the role of lipin-3?and the interaction between lipin-1 and lipin-3?in adipocyte function are likely to be relevant to human adipose tissue biology, since the balance between lipin-1 and lipin-3 may influence the differential susceptibility of lipin-1-deficient mice and humans to overt lipodystrophy. A third discovery is that lipin-2 and lipin-3 each contribute to PAP activity in the small intestine, and that the loss of both proteins leads to a dramatic accumulation of cytosolic TAG droplets in enterocytes. We propose that lipin-2 and lipin-3 PAP activities are important for intestinal lipid homeostasis and chylomicron assembly. The implication of lipins in this process is novel given that intestinal TAG synthesis has been attributed primarily to the monoacylglycerol pathway. Our studies will shed new light on a fundamental process of lipid metabolism in the small intestine. .